Bisphenol A (BPA) is an endocrine disrupting chemical causing ubiquitous exposures in the developed world. Both animal studies and recent epidemiological evidence indicate that BPA targets the male reproductive tracl<, producing testicular injury and alterations in sperm parameters. Standard GLP-based approaches in rodent models have been relatively insensitive in detecting BPAinduced abnormalities in the male reproductive tract. We propose the use of well-established morphometric assessments ofthe testis, including quantitation of retained spermatid heads and the incidence of TUNELpositive germ cells, to enhance the sensitivity of detecting testicular effects of BPA exposure. In addition, we will identify molecular biomarkers of effect in the testis and sperm, by examining global mRNA and miRNA levels and genome-wide DNA methylation. Measuring these molecular biomarkers in sperm is expected to be particularly sensitive and specific, since sperm are an homogeneous population of cells that developed throughout their life history within the seminiferous epithelium. The GLP design provides multiple doses, and a positive estradiol control, allowing robust testing for the presence of a BPA-induced effect. In addition, we will assess both the paternal FO rats and their progeny, addressing differences across generations. The goal of this project is to test the following working hypothesis: addition ofthe enhanced morphological and molecular endpoints with the GLP design will provide more sensitive and specific biomarkers of BPA-induced male reproductive toxicity. The GLP study will be enhanced by morphometric assessments and molecular biomarkers of effects through the following Specific Aims: Specific Aim #1: will enhance the assessment of BPA-induced testicular injury using advanced histological and morphometric endpoints. Specific Aim #2: characterize the caudal sperm and testis mRNA and miRNA transcript content. Specific Aim #3: develop the caudal sperm and testis methylome.